This project will be concerned with the properties of the regulatory system which controls the activity of the adenylate associated with a particulate fraction isolated from brain tissue. This membrane bound enzyme catalyzes the formation of cyclic 3', 5'-AMP. The enzyme of brain appears to be associated with receptors for various neurotransmitters, and stimulation by these compounds results in increased levels of the cyclic nucleotide in brain tissue. The details of the mechanism by which the activity of this enzyme is regulated have not been determined. Recent evidence obtained in this laboratory indicates that it is possible to investigate the properties of this regulatory system in isolated preparations of brain adenylate cyclase. A working hypothesis based upon these data proposes that the adenylate cylase of brain can exist in active and inactive forms. The two forms of the enzyme are interconvertible through separate reactions. Activation appears to require divalent cations and may require ATP or some other nucleoside triphosphate. Inactivation appears to be inhibited by fluoride ions, a well known modifier of adenylate cyclase activity. The experiments described in this proposal are designed to test the validity of this hypothesis and to investigate the properties of the regulatory system. The nature of the reactions which produce activation and inactivation will be determined and experiments will be performed to determine if they are catalyzed by specific enzymes. If so, then the properties of these enzymes will be investigated. The possibility that this regulatory system is involved in the response of brain adenylate cyclase to neurohormones will be investigated. It is anticipated that the results of these investigations will lead to a better understanding of the mechanism by which the cell regulates the rate of formation of cyclic 3',5'-AMP.